Project summary Currently 72% of women who practice contraception use hormonal methods, but there is frequent dissatisfaction with these methods, due to quality of life and safety concerns. Therefore, there is a clear need for new approaches to non-hormonal female contraceptives that are easy to use, woman-applied, and have a controllable duration of action. Reversible immunocontraception offers a non-hormonal solution, where anti- sperm antibodies are introduced into the female reproductive tract (FRT) and inhibit sperm function. This approach, though, has a number of challenges including: discovery of a specific and effective monoclonal antibody (mAb) against a human sperm antigen, and a safe and reliable method for introduction of mAbs that is temporally and spatially controllable. For this grant application, reproductive biology expertise and a molecular toolbox has been brought together to overcome these challenges. An anti-sperm antibody has been identified with well-characterized mechanisms of action that impact fertility, and an innovative method has been identified to deliver the antibody to the FRT. Here a synthetic mRNA-based approach is proposed to deliver a sperm agglutinating and mucus trapping antibody to the FRT. To date, the ability to express high levels of antibody for over 28 days with a single administration of mRNA in the FRT of sheep has been demonstrated. Delivery is achieved through direct, rapid, aerosol exposure of the FRT epithelium to naked mRNA in water. Persistence of the antibody in secretions was achieved through the incorporation of a GPI-linker into the heavy chain of the antibody. This combination allows for rapid delivery and expression, and for controllable persistence of the protective mAb in secretions. In the R61 phase, the mechanism of delivery under relevant conditions will be optimized and explored; antibody design questions will also be addressed that will allow for tunable persistence of the antibody in mucus secretions. In the R33 phase, pharmacokinetic experiments will be performed in macaques, as well as continued antibody optimization. Last, sperm challenge experiments will be performed in vivo to demonstrate efficacy. The long-term goal is to develop a cost-effective mRNA-based approach for expressing anti-sperm antibodies in the FRT. The short-term goals are to optimize the approach, answer mechanistic questions, and demonstrate pre-clinical feasibility in the macaque model. If successful, a new paradigm for contraception will be demonstrated, one that can be combined with the co-expression of anti-STI antibodies, such as those against HIV and HSV-2.